In mammals, activin and inhibin are important regulators of FSH secretion. Previous studies have demonstrated that primary ovine pituitary cells express different activin receptor subtypes: activin receptor-like (ALK)2, ALK4, activin type II receptor A (ActRIIA), ActRIIB and Smad proteins in vitro. Here, we have carried out physiological studies to investigate the pattern of mRNA expression of the activin receptor subunits in the ewe pituitary throughout the oestrous cycle. The oestrous cycles of ewes were synchronized with progestagen sponges. The animals were killed 36 h (before the preovulatory surge, n=4), 48 h (during the preovulatory surge, n=4), 72 h (during the second surge of FSH, n=6) and 192 h (during the luteal phase, n=4) after sponge removal. Using Northern blots, we have shown that the levels of ALK2, ALK4 and ActRIIB mRNA were significantly higher before the preovulatory surge and during the secondary surge of FSH as compared with both during the preovulatory surge and the luteal phase, whereas the level of the ActRIIA mRNA was similar throughout the oestrous cycle. Using Western blots we have also demonstrated that the level of phosphoSmad2 did not vary during the reproductive cycle. Inhibin binding protein (InhBP/p120) and the transforming growth factor-type III receptor, betaglycan, have been identified as putative inhibin co-receptors. In this study, we cloned a fragment of both InhBP/p120 and betaglycan cDNAs in the ewe and showed by Northern blot that pituitary betaglycan and InhBP/p120 mRNA levels did not fluctuate across the oestrous cycle nor did they correlate with serum FSH levels.

The complete coding sequence for the bovine thyrotropin (TSH) receptor was derived using a modified PCR cloning strategy. The bovine thyrotropin receptor conforms to the pattern of receptor interacting with membrane-bound G-protein already established in other species for TSH and gonadotropins receptors. The cDNA for the bovine TSH receptor consists of an open reading frame 2289 nucleotides in length, corresponding to a protein of 763 amino acids (estimated molecular mass of 86.4 kDa) which includes a 20 amino acid putative leading signal peptide. The receptor consists of a large NH2-terminal extracellular membrane domain of 417 amino acids with 5 potential N-linked glycosylation sites, a transmembrane domain (265 amino acids) consisting of 7 putative membrane alpha-helix spanning segments, and an intracytoplasmic COOH-terminal domain (82 amino acids). The bovine TSH receptor is one amino acid less than the corresponding sequence in dog, human, rat and mouse. Cysteine residues (n = 22) were conserved when compared with other TSH receptors. Three potential phosphorylation sites were found in the transmembrane domain and the COOH-terminal domain. As with other members of this receptor family, alternative splicing was observed. A transcribed but truncated TSH receptor of 1769 nucleotides was demonstrated, lacking half of the V segment of the transmembrane domain up to the COOH-terminal domain of the full length TSH receptor. Additionally, alternative transcriptional start sites were observed. Northern blot analysis using a probe (1170 bp) spanning part of the extracellular domain up to the first loop of the transmembrane domain showed specific expression in the bovine thyroid gland with major transcripts of 9.3 and 4.3 kb, and a minor transcript of 3.8 kb being detected.